Numerous vascular conditions afflict mammals. Such conditions include, but are not limited to, coronary and peripheral arterial diseases, chronic rejection, vasculopathy associated with diabetes, pulmonary vascular conditions (e.g., pulmonary arterial hypertension and chronic obstructive pulmonary disease), ocular vascular conditions (e.g., intraocular pressure and glaucoma), sexual dysfunction vascular conditions (e.g., erectile dysfunction and vulvodynia), and dermal vascular conditions (e.g., Raynaud's phenomenon, scleroderma and wound healing).
A known compound that has a general dilatory effect on the vascular system is nitric oxide (NO). See Nathan et al., Cell (1994) 78:915-916. NO plays an essential role in mammalian physiology and is responsible for various functions including vascular tone, endothelium dependent reactions, activation of soluble guanylate cyclase, neurotransmission in the central and peripheral nervous systems, and activated macrophage cytotoxicity. In particular, as the endothelial-derived relaxation factor (EDRF), NO plays a crucial role in vasodilation throughout the body and is a known antagonist of endothelin-1, one of the most potent mammalian vasoconstrictors. See Palmer, R. M., et al., Nature (1987) 327:524-526. It is believed that NO functions by binding to heme and activating soluble guanylate cyclase to increase the cellular content of cGMP and activate cGMP-dependent protein kinases. The latter both generate vasodilatory effects and reduce blood vessel tone. Other functions of NO include the inhibition of platelet adherence and aggregation, and the inhibition of vascular smooth muscle proliferation and leukocyte adherence. Thus, NO is considered an inhibitor of stenosis, restenosis, vascular inflammation, vascular cell proliferation, thrombosis, atherosclerosis, and arteriosclerosis.
NO is synthesized, at least in part, from L-arginine by a family of enzymes known as nitric oxide synthases (NOS). It is believed that NOS converts L-arginine, NADPH, and oxygen into citrulline, NADH, and NO. NOS occur in several isoforms: an endothelial nitric oxide synthase (eNOS), a machrophage or inducible nitric oxide synthase (iNOS), and a neuronal nitric oxide synthase (nNOS). Unlike its name, eNOS has been detected not only in endothelial cells and blood vessels, but also in epithelium of tissues including, but not limited to, bronchial cells and neurons of the brain, especially in the pyramidal cells of the hippocampus. Furthermore, iNOS has been detected not only in macrophages but also in cells such as hepatocytes, chrondrocytes, endothelial cells, and fibroblasts, in particular under conditions of endothelial damage or as part of a response to injury.
The NOS isoforms can also be categorized as either constitutive or inducible. Constitutive NOS (cNOS) include eNOS and nNOS, while iNOS is inducible. cNOS are usually present in a cell, but remain inactive until intracellular calcium levels increase resulting in enhanced calcium/calmodulin binding and subsequent activation. Unlike cNOS, iNOS is calcium independent and is not normally present in cells. However, iNOS can be induced by lipopolysaccharides and certain cytokines. It is postulated that cytokine activity affects gene expression/splicing, mRNA stability, and protein synthesis, resulting in iNOS. It is also expected that the induced form of NOS produces a much greater amount of NO than cNOS, and may even result in toxicity when the L-arginine supply is limited. Induction of iNOS can be inhibited by gluococorticoids and some cytokines.
Recent studies suggest that NOS inhibitors may be associated with endothelial vasodilator dysfunction. In particular, asymmetric dimethylarginine (ADMA), and to a lesser extent, N-monomethylarginine (NMA) are associated with endothelial vasodilator dysfunction. Patients with coronary and peripheral arterial disease and those with renal failure have greater amounts of plasma ADMA. However, it has been shown that while exogenous ADMA vasoconstricts vascular rings in vitro, the vasoconstriction effect can be reversed by L-arginine.
Formation of NO by eNOS is thought to play an important role in normal blood pressure regulation, prevention of endothelial dysfunction such as hyperlipodemia, arteriosclerosis, atherosclerosis, thrombosis, restenosis, ischemia, and apoptosis. eNOS is the predominant synthase present in brain and endothelium and may be active under basal conditions. Yamada M., J. Cereb. Blood Flow Metab. (2000) April; 20(4):709-17. eNOS can be stimulated by increases in intracellular calcium that occur in response to receptor-mediated agonists or calcium ionophores. Studies further suggest that cNOS activity can be regulated by a negative feedback manner by NO.
Since intracellular levels of L-arginine are normally greater that NOS enzyme, NO syntheses generally do not depend on extracellular supplementation. See Harrison, D. G., et al. J. Clin. Invest. (1997) 100:2153-2157. However, under certain circumstances, local L-arginine concentrations might become rate limiting. Such circumstances might include local tissue inflammation, elevated plasma or tissue levels of ADMA; inflammation-induced expression of the iNOS; increased expression of arginase, and presence of iNOS stimulants such as IFN-.gamma. and LPS. See Guoyao, et al. Biochem. J. (1998) 366:1-17.
As a free radical gas, NO has an extremely short half-life. See Morris et al., Am. J. Physiol. (1994) 266:E829-E839. Thus, it is desired to increase the effective amount of NO in a cell, tissue, and/or organ in order to induce vascular relaxation, dilation, or vascularization, and oxygenation, and other NO mediated biological processes. Previous publications suggest that NO can be increased by administering to an organism a NO donor that releases NO, e.g., glyceryl trinitrate, isosorbide 5-mononitrate, isosorbide dinitrate, pentaerythritol, pentaerythritol tetranitrate, etc. One of the greatest limitations in administering a NO donor is that in vivo administration of such compounds can induce severe systemic hypotension. See Heros et al., Surgical Neurology, (1976) 5:354-362. Others have suggested that L-arginine monomers can be administered to prevent vasoconstriction and vascular conditions such as atherosclerosis and restenosis. See Cooke et al., U.S. Pat. No. 5,428,070. L-arginine is taken up by cells by way of the y+ transporter. This transport mechanism is limited according to the expression of the transporter and other molecules competing for the transporter (including ADMA).
Therefore, it is desirable to find new compositions and methods for treating and/or preventing vascular conditions or to increase the local tissue concentration of NO without causing clinically significant systemic hypotension.